The major aims of the proposed research is to more fully define the mechanisms by which CA2+ regulates cell function during such processes as the hormonal and ionic regulations of aldosterone secretion, from adrenal glomerulosa cells. Cell proliferation of cultured 3T3 cells, and leukocytes function. Specific aims include an elucidation of the temporal patterns of protein phosphorylation, inositol lipid metabolism, Ca2+ metabolism, and protein kinase C function during angiotensin II and bombesin-mediated response in, respectively, glomerulosa and 3T3 cells. The role of type II CaM-dependent protein kinase in the initial phase of cellular response; the immunocytochemical localization of the C-kinase and the possible role of M-kinase in sustained cellular response; the mechanism by which information is conveyed from cell surface to cell interior via the C-kinase branch of the Ca2+ messenger system during the sustained phase of cellular response focusing particularly on changes in Na+/H+ exchange and possible protein kinase cascades; the molecular basis of memory in the C-kinase-mediated phase of cellular response; and a reexamination of the temporal pattern of events in the cAMP messenger system during ACTH-mediate aldosterone secretion from adrenal glomerulosa cells as the system in which to explore these issues. Methods will include studies of Ca2+ fluxes and measurements of intracellular free Ca2+ using both fura 2 and aequorin as indicators; studies of the turnover of inositol lipids and inositols phosphates by both chemical and radio- isotope methods; 2-D gel electrophoresis of the phosphoproteins obtained from extracts of cells; and both light and EM immunocytochemical localization of C-kinase. The problem under investigation is of broad biological interest, and has implications for a wide range of biological phenomena. A better understanding of Ca2+ messenger system function is likely to alter our views of cell growth and proliferation, peptide and steroid hormone secretion, and smooth muscle contraction.